title: Evaluation of PMS-PCR technology for detection of Mycobacterium avium subsp. paratuberculosis directly from bovine fecal specimens.
abstract: Mycobacterium avium subsp. paratuberculosis (MAP) causes paratuberculosis, or Johne's disease, in animals. Diagnosis of MAP infection is challenging because of the pathogen's fastidious in vitro growth requirements and low-level intermittent shedding in feces during the preclinical phase of the infection. Detection of these "low-shedders" is important for effective control of paratuberculosis as these animals serve as sources of infection for susceptible calves. Magnetic separation technology, used in combination with culture or molecular methods for the isolation and detection of pathogenic bacteria, enhances the analytical sensitivity and specificity of detection methods. The aim of the present study was to evaluate peptide-mediated magnetic separation (PMS) capture technology coupled with IS900 PCR using the Roche real-time PCR system (PMS-PCR), in comparison with fecal culture using BACTEC-MGIT 960 system, for detection of MAP in bovine fecal samples. Among the 351 fecal samples 74.9% (263/351) were PMS-PCR positive while only 12.3% (43/351) were MGIT culture-positive (p=0.0001). All 43 MGIT culture-positive samples were also positive by PMS-PCR. Mean PMS-PCR crossing-point (Cp) values for the 13 fecal samples with the highest number of MAP, based on time to detection, (26.3) were significantly lower than for the 17 fecal samples with <100 MAP per 2g feces (30.06) (p<0.05). PMS-PCR technology provided results in a shorter time and yielded a higher number of positive results than MGIT culture. Earlier and faster detection of animals shedding MAP by PMS-PCR should significantly strengthen control efforts for MAP-infected cattle herds by helping to limit infection transmission at earlier stages of the infection.
authors.affiliation: Biochemistry and Microbiology Department, Faculty of Sciences, Universidad Austral de Chile, PO Box 167, Valdivia, Chile. Electronic address: miguelsalgado@uach.cl.
authors.forename: M
authors.initials: M
authors.lastname: Salgado
authors.forename: P
authors.initials: P
authors.lastname: Steuer
authors.forename: E
authors.initials: E
authors.lastname: Troncoso
authors.forename: M T
authors.initials: MT
authors.lastname: Collins
mesh_terms: D000818:Animals; D001431:Bacteriological Techniques / Q000379:methods / Q000662:veterinary*; D002417:Cattle; D002418:Cattle Diseases / Q000175:diagnosis* / Q000382:microbiology; D005243:Feces / Q000382:microbiology*; D060328:Magnetic Phenomena; D016927:Mycobacterium avium subsp. paratuberculosis / Q000235:genetics* / Q000302:isolation & purification; D010283:Paratuberculosis / Q000175:diagnosis* / Q000382:microbiology; D016133:Polymerase Chain Reaction / Q000592:standards / Q000662:veterinary*; D012680:Sensitivity and Specificity
keywords: Diagnose; Liquid culture; Mycobacterium avium subsp. paratuberculosis; PMS–PCR; Paratuberculosis